崑嵛林蛙（Rana kunyuensis）作为林蛙属的一种，仅自然生活于中国山东省烟台市昆嵛山境内。在本研究中，从合成的崑嵛林蛙皮肤cDNA文库中克隆获得一条279 bp的cDNA序列，该序列编码一种新型抗菌肽分子，命名为amurin-9KY。Amurin-9KY前体由62个氨基酸残基组成，其中成熟肽由14个氨基酸残基组成，包括由2个半胱氨酸残基形成的分子内七肽环和酰胺化的C-末端。这些结构特点表明amurin-9KY是一种新型两栖类抗菌肽家族成员之一。尽管amurin-9KY和之前从黑龙江林蛙（Rana amurensis）中发现的amurin-9AM具有较高的相似性，但是到目前为止对于amurin-9家族抗菌肽的结构和功能仍然未知。在本论文中，我们设计合成了amurin-9KY和3个改造体多肽amurin-9KY1-3，研究了C-末端酰胺化和七肽环对amurin-9KY结构和功能的影响。结果表明C-末端酰胺化对amurin-9KY的抗菌活性十分重要，而C-末端酰胺化和七肽环对amurin-9KY的溶血活性同时具有影响。圆二色谱（Circular dichroism, CD）结果表明上述4种多肽在TFE/H2O (v/v 1:1)溶液中均为α-螺旋型结构，而在水溶液中为无规则结构。打断七肽环的二硫键使2个半胱氨酸的巯基游离能够显著增加amurin-9KY的抗氧化活性。扫描电子显微镜（Scanning electron microscopy, SEM）同时也被用于研究amurin-9KY的抗菌机理。
Rana kunyuensis is a species of brown frog that lives exclusively on Kunyu Mountain, Yantai, China. In the current study, a 279-bp cDNA sequence encoding a novel antimicrobial peptide (AMP), designated as amurin-9KY, was cloned from synthesized double-strand skin cDNA of R. kunyuensis. The amurin-9KY precursor was composed of 62 amino acid (aa) residues, whereas the mature peptide was composed of 14 aa and contained two cysteines forming a C-terminal heptapeptide ring (Rana box domain) and an amidated C-terminus. These structural characters represent a novel amphibian AMP family. Although amurin-9KY exhibited high similarity to the already identified amurin-9AM from R. amurensis, little is known about the structures and activities of amurin-9 family AMPs so far. Therefore, amurin-9KY and its three derivatives (amurin-9KY1–3) were designed and synthesized. The structures and activities were examined to evaluate the influence of C-terminal amidation and the heptapeptide ring on the activities and structure of amurin-9KY. Results indicated that C-terminal amidation was essential for antimicrobial activity, whereas both C-terminal amidation and the heptapeptide ring played roles in the low hemolytic activity. Circular dichroism (CD) spectra showed that the four peptides adopted an a-helical conformation in THF/H2O (v/v 1:1) solution, but a random coil in aqueous solution. Elimination of the C-terminal heptapeptide ring generated two free cysteine residues with unpaired thiol groups, which greatly increased the concentration-dependent anti-oxidant activity. Scanning electron microscopy (SEM) was also performed to determine the possible bactericidal mechanisms.